Particle detection systems and methods for on-axis particle detection and/or differential detection

ABSTRACT

Provided herein are optical systems and methods for detecting and characterizing particles. Systems and method are provided which increase the sensitivity of an optical particle counter and allow for detection of smaller particles while analyzing a larger fluid volume. The described systems and methods allow for sensitive and accurate detection and size characterization of nanoscale particles (e.g., less than 50 nm, optionally less than 20 nm, optionally less than 10 nm) for large volumes of analyzed fluids.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of and priority to U.S. ProvisionalPatent Application No. 62/838,835, filed Apr. 25, 2019, which is herebyincorporated by reference in its entirety.

BACKGROUND OF INVENTION

Advancement of technologies requiring cleanroom conditions has resultedin a need for the detection and characterization of smaller and smallerparticles. For example, microelectronic foundries pursue detection ofparticles less than 20 nm in size, and in some cases less than 10 nm insize, as they may affect the increasingly sensitive manufacturingprocesses and products. Similarly, the need for aseptic processingconditions for manufacturing of pharmaceuticals and biomaterialsrequires accurate characterization of viable and non-viable particles toaddress compliance standard relating to health and human safety.

Typically, these industries rely on optical particle counters fordetection and characterization of small particles. The ability to detectsmaller particles requires new approaches for optical particle countingsuch as systems employing increasing laser powers, shorter excitationwavelengths and more complex techniques such as condensation nucleicounting, which in turn can dramatically increase the cost and overallcomplexity of devices for detection of nanometer scale particles. Thesenew approaches can also require more frequent calibration andmaintenance to provide the necessary reliability and reproducibility.

Various optical particle counters are known in the art, for example,scattered light optical particle counters are provided in U.S. Pat. No.7,916,293 and transmission/extinction particle counters, including thoseutilizing structured beams and/or interferometry are provided in U.S.Pat. Nos. 7,746,469, 9,983,113, 10,416,069, US Patent Publication Nos.2019/0277745 and US 20170176312, and PCT international Publication WO2019/082186. Each of these references are hereby incorporated in theirentirety and specifically to illustrate particle counter systemcomponents and configurations that are useful for the detection andcharacterization of small particles.

It can be seen from the foregoing that there is a need in the art forsystems and methods that provide enhanced optically sensing particleshaving small size dimensions.

SUMMARY OF THE INVENTION

Provided herein is are optical particle counter and/or analyzer systemsand methods which increase the sensitivity, accuracy and throughput ofan optical particle counter to allow for detection and characterizationof smaller particles over large volumes of sampled fluid, for example,using on-axis detection of transmitted and/or forwarded scattered light,optionally using a probe beam comprising a structured beam such as adark beam and optionally using a differential detection system. In someembodiments, the present systems and methods provide for detection andcharacterization of particles in fluids using a structured beam, such asa dark beam, incorporating optical geometries, detector configurationsand signal analysis techniques allowing for an enhancement in the amountof sample fluid analyzed as a function of time, an increase in overallparticle detection sensitivity for small particles (e.g., effectivelateral dimensions (e.g., diameter) of 10 microns, or optionally 1microns or optionally 500 nanometers) and/or a suppression of falsepositive indications.

The present systems and methods are particularly well-suited forparticle measurement (e.g., detection and/or size characterization)using on-axis particle measurements by detection of transmitted andforward scattered light and/or differential detection configurations andmethods. The present systems and methods are highly versatile and may beimplemented using a range of particle measurement techniques including:(i) particle detection using interferometric detection of particles;(ii) particle detection using Gaussian and non-Gaussian beam, such astructured beam, dark beam, etc. (iii) particle detection usingdifferential detection, (iv) particle detection using multipasstechniques (e.g., dual pass) and/or (v) particle detection usingpolarization control.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, and optionally a differential detection configuration, withoptical and/or flow cell elements (e.g., translators, oscillators,piezoelectric elements, etc.) providing for rapid translation (e.g.,faster than the average velocity of the particle as it passes throughthe beam) of the flow cell along a lateral direction (e.g., along anaxis orthogonal to the beam axis) and/or along the z-axis (e.g., alongan axis along the beam axis between the optical source and detector) toachieve greater sampled volume of fluid per unit time relative tosystems not employing translation. In an embodiment, for example, thelaser source or flow cell is translated, for example via a translator oroscillator, to as to increase the volume of sample fluid analyzed perunit time via particle detection using a structured beam such as a darkbeam. In an embodiment, for example, the flow cell is translated at anaverage translation velocity that is at least two times faster than theaverage particle velocity as it travels through the beam and optionallyfor some applications the flow cell is translated at an averagetranslation velocity that is 5-100× faster than the average particlevelocity as it travels through the beam. In some embodiments, forexample, the flow velocity of the fluid containing particle is selectedover the range of 5-300 cm/sec. In some embodiments, for example, theflow cell is translated at an average translation velocity greater thanor equal to 25 cm/sec. In an embodiment, the translator providestranslation of the flow cell along a distance selected from the range of10-1000 microns. In some embodiments, the translator undergoes periodictranslation at a frequency selected from the range of 100 kHz-100 MHz.Translation may be a periodic translation in one direction or more thanone direction, including an oscillation and/or may a linear translationor a non-linear translation. In some embodiments, for example, theoscillator oscillates at a frequency high enough such that the flow cellcompletes its displacement in less time than is required for theparticles to pass through the area of high radiation density. In someembodiments, the translator is an oscillator that oscillates at afrequency selected from the range of 100 kHz 100 MHz.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, and optionally a differential detection configuration, withuse of pixelated photodetector(s) having pixel areas corresponding to(e.g., within a factor of 1.5 and optionally a factor of 1.2) spatialextent of the particle beam interaction signal within the beam, therebyproviding an enhancement for detection of more particle transitionsleading to an improvement of the sample volume analyzed per unit time.In some embodiments, such pixelated detection configurations allow forbreaking out of particle signal image from the surrounding “larger” beamimage. In some embodiments, the systems and methods incorporate apixelated photodetector with one or more rows of elements so as toprovide for efficient differential detection between at least twoelements. Pixel widths each independently selected from the range of 10to 500 microns are useful in certain embodiments and optionally for someapplications each independently selected from the range of 50-100microns.

In some embodiments, systems and methods of the invention combineon-axis particle measurements by detection of transmitted and forwardscattered light, optionally using a differential detection configurationand/or using a structured beam such as a dark beam, with additionaloff-axis detection of scattered light or fluorescence for distinguishingbetween biological (e.g. microbial particles) and non-biologicalparticles. Biological particles, such as microbes, are composedprimarily of water, so they have a very low index of refraction contrastwith water and hence scatter very little. Accordingly, biologicalparticles generate a small side scattering signal or are not detectedusing side scattering. Biological particles do create a strong responsesignal in the present methods and systems using on-axis detection oftransmitted light and forward scattered light, for example usingstructure beam and/or differential detection techniques. In some methodsand systems of the invention, observation of (or comparison of) a largeon-axis signal compared to the side scatter signal or observation of asignal at the on-axis detector without a corresponding signal at theside scatter detector is used to characterize a particle as a biologicalparticle, such as a microbial particle. Alternatively, in some methodsand systems of the invention, observation of (or comparison of)comparable on-axis signal and side scatter signal or observation of bothsignals at the on-axis detector and at the side scatter detector is usedto characterize a particle as a non-biological particle.

In some embodiments, systems and methods of the invention use on-axisparticle detection by detection of transmitted and forward scatteredlight using a structured beam, such as a dark beam, to characterize therefractive index of the particle, for example, relative to (e.g.,greater or less than) the refractive index of the media the particle isin (e.g., the composition of the fluid flow). This aspect allows fordetermining attributes of the composition of the particle, for example,providing a means for distinguishing between metallic and non-metallicparticles based on refractive index. In systems incorporating adifferential detection configuration, for example, reliable andrepeatable flipping of the “classical” particle signal may be used foraccurately distinguishing between metallic and non-metallic particles.In some embodiments, for example, difference in the refractive index ofthe particle relative to the refractive index of the carrier fluidresults in a signal that may be used to characterize the opticalproperties and composition of the particles.

If the analyzed particle has a refractive index higher than therefractive index of the carrier fluid, for example as in the case ofpolystyrene latex (PSL) particles in water media, a bright fringe isobserved at the top of the beam and a dark fringe is observed at thebottom of the beam for conditions wherein the particle enters the beamin the flow cell. For example, when the particle enters the beam fromthe bottom. On the other hand, if the analyzed particle has a refractiveindex lower than the refractive index of the carrier fluid, as in thecase of gold nanoparticles in water media, the fringe pattern isreversed for conditions of wherein the particle enters the beam in theflow cell, such as when the particle enters the beam from the bottom,such that a dark fringe is observed at the top of the beam and a brightfringe is observed at the bottom of the beam. Therefore, by observingand characterizing the sequence, order and/or positioning of the brightfringe and the dark fringe during the particle interaction with the beamthe refractive index relative to the carrier (e.g., greater than or lessthan) of the particles can be characterized and, therefore, informationrelating to composition of the particles can also be inferred.Differential detection provides an efficient and accurate means ofcharacterizing the sequence, order and/or position of dark fringe andbright fringe in the present systems and techniques as a function oftime during the trajectory of the particle through the beam, forexample, at times (i) when the particle first enters the beam (e.g.,from the bottom), (ii) when the particle passes through the beam waistand (iii) when the particle exists the beam.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, optionally using a differential detectionconfiguration and/or using a structured beam such as a dark beam, withan adjuster to balance the differential detector, such as first andsecond detector regions corresponding to a top pixel or top subset ofpixels and a bottom pixel or subset of pixels, across the beam, forexample, by moving the detector using a positioner and/or actuating amirror before the detector to balance signal between first and seconddetector regions corresponding to a top pixel or top subset of pixelsand a bottom pixel or subset of pixels. In an embodiment of thisdifferential detection aspect, adjustment of the detector and/or thebeam position on the detector using an adjustor provides for enhancednoise cancellation, particularly for circumstances wherein the positionof the beam is susceptible to vibration and acoustical inputs, forexample, when the system is not isolated from source of such vibrationand acoustical input. In an embodiment of this differential detectionaspect, a portion of the beam is provided to an imager or multipledetector (e.g., quad detector) to provide feedback for beam powerdensity, spot size in flow cell. In some embodiments, the adjustor isoperated via closed loop control, for example, by periodic measurementof the noise amplitude of the differential signal when particles are notpresent and active adjustment of the differential detector positionand/or beam position on the differential detector so as to the noiseamplitude of the differential signal.

In some embodiments, the present systems and methods include one or moreadjustors to ensure that the laser beam intensity is balanced betweenthe upper half and the lower half of a differential detector, forexample, so as to reduce noise and enhance signal In an embodiment, forexample, a closed loop system is employed that that determines andanalyzes the noise amplitude of the differential signal when particlesare not present. In an embodiment, a steering mirror is used to adjustthe beam position on the detector to minimize noise levels of thedifferential signal. This condition occurs when the beam power isuniformly split between the upper and lower elements, such as uniform towithin 20% and optionally uniform to within 10%. Similarly, such controlcan also be achieved through translating the detector position androtating the detector to align the beam and detector axes.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, with an optical modulator with a lock-in amplifier; inconjunction with cooled detector for improved signal-to-noise ratio.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, and optionally a differential detection configuration, with acollimating or imaging the beam onto the photodetector.

In some embodiments, for example, systems and methods of the inventioncombine on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, and optionally a differential detection configuration, with aknife edge prism to split the beam onto separate photodetectors

In an aspect, a particle detection system comprises a flow cell forflowing a fluid containing particles; an optical source for generatingone or more coherent beams of electromagnetic radiation; a beam shapingsystem for passing the one or more coherent beams of electromagneticradiation through the flow cell, thereby generating electromagneticradiation scattered by the particles; at least one optical detectorarray for receiving electromagnetic radiation from the flow cell,wherein the optical source, beam shaping system and optical detectorarray are configured to allow for detection of the particles. In anembodiment, the optical source, beam shaping system and optical detectorarray are configured to provide interferometric detection of particles.In an embodiment, the optical source, beam shaping system and opticaldetector array are configured to provide structured beam detection ofthe particles by passing a structured probe beam of coherentelectromagnetic radiation through the flow cell. In an embodiment, theoptical source and optical detector array are configured to providestructured dark beam detection of the particles, such as a structureddark beam characterized by a spatial intensity profile having region ofattenuated intensity, such as a centerline decrease in intensity.

In an embodiment, the optical detector array is positioned in opticalcommunication with the flow cell for receiving incident electromagneticradiation transmitted through the flow cell and electromagneticradiation scattered by the particle, for example, wherein theelectromagnetic radiation scattered by the particle comprises forwardscattered electromagnetic radiation. In an embodiment, the incidentelectromagnetic radiation transmitted through the flow cell and theelectromagnetic radiation scattered by the particle undergo constructiveand/or destructive optical interference, for example, thereby generatingone or more diffraction patterns. In an embodiment, the optical detectorarray is provided at a scattering angle that is within 5 degrees of zerodegrees relative to the optical axis of the incident beam, optionallyfor some applications at a scattering angle that is within 1 degree ofzero degrees relative to the optical axis of the incident beam,optionally for some applications at a scattering angle that is within0.5 degree of zero degrees relative to the optical axis of the incidentbeam, and optionally for some applications at a scattering angle that iswithin 0.1 degree of zero degrees relative to the optical axis of theincident beam. In an embodiment, the optical detector array is providedin optical communication with the flow cell for detecting theinteraction of the particle and the electromagnetic radiation scatteredby the particle with the illuminating wave front.

In an embodiment, the source provides a coherent incident beam to theflow cell, such as a Gaussian incident beam. System and methods of theinvention are also well-adapted for structured beam detection using astructured beam such as dark beam. In an embodiment, the optical sourcecomprises one or more shaping and/or combining optical elements forgenerating the one or more coherent beams of electromagnetic radiation.In an embodiment, the one or more shaping and/or combining opticalelements are diffractive elements, polarizing elements, intensitymodulating elements, phase modulating elements or any combination ofthese. In an embodiment, the one or more coherent beams ofelectromagnetic radiation comprises a structured, non-Gaussian beam. Inan embodiment, the one or more coherent beams of electromagneticradiation comprises a dark beam. In an embodiment, the one or morecoherent beams of electromagnetic radiation comprises a beamcharacterized by one or more line singularities. In an embodiment, theone or more coherent beams of electromagnetic radiation comprises ananamorphic beam. In an embodiment, the one or more coherent beams ofelectromagnetic radiation comprises an anamorphic beam in a top hatconfiguration.

The systems and methods are compatible with a wide range of detectorsand detector configurations. In an embodiment, forward looking on axisdetector pair(s) are provided, for example at a scattering angle that iswithin 20 degrees of zero degrees relative to the optical axis of theincident beam, optionally for some applications a scattering angle thatis within 5 degrees of zero degrees relative to the optical axis of theincident beam, optionally for some applications at a scattering anglethat is within 1 degree of zero degrees relative to the optical axis ofthe incident beam, optionally for some applications at a scatteringangle that is within 0.5 degree of zero degrees relative to the opticalaxis of the incident beam, and optionally for some applications at ascattering angle that is within 0.1 degree of zero degrees relative tothe optical axis of the incident beam. Differential detection may beused in the present systems and methods to provide a significantreduction of noise, for example by using a detector configuration withfirst and second active regions aligned to receive portions of theincident beam, optionally wherein the beam power is uniformly splitbetween the first and second active regions.

The systems and methods of the invention provide detection of particlesin flowing fluids, including detection, counting and sizing of singleparticles in a fluid flow. In an embodiment, the fluid is a liquid or agas. In an embodiment, the system is for detection of particles inliquid chemicals. In an embodiment, the system is for detection ofparticles in ultrapure water. In an embodiment, the system is fordetection of particles in high pressure gases. In an embodiment, thesystem is for detection of particles in air. In an embodiment, thesystem is for detection of particles on surfaces.

In some embodiments, the present systems and methods are for analyzinglarge sample volumes per unit time by: (i) adjusting the depth of focusof the beam of electromagnetic radiation, (ii) increasing the effectivescanning area of the beam and/or (iii) increasing the signal-to-noiseratio generated by the detector elements of the particle counter. Thedescribed systems and methods may allow for detection of nanoscaleparticles (e.g., less than 50 nm, optionally less than 20 nm, optionallyless than 10 nm), for example using lower laser power requirements thanin conventional optical particle counters.

One method of increasing the effective scanning area of a beam or laserof an optical particle counter is to rapidly translate the beam throughthe target flow cell, such that the beam effectively scans a largercross-sectional area or volume of the fluid being analyzed. The beam maybe translated by various methods, including oscillating the flow cell,the optical focusing system or the electromagnetic source. Theoscillation may be at a higher frequency than the transit time ofparticles within the fluid (based on the flow rate of the fluid in theflow cell) reducing or elimination the possibility that particles couldbe missed by beam due to the movement. The oscillation may be in thex-direction (laterally with regard to the beam propagating in thez-direction) in the y-direction (vertically with regard to the beampropagating in the z-direction), and/or in the z-direction (along thebeam path). Oscillators may be various acoustic, electric or mechanicaldevices known in the art, for example, a piezoelectric device.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation to generate an area of high radiation density,such as a focused beam region, within the flow cell; iv) an translator,such as an oscillator, operably connected to the flow cell fortranslating the flow cell closer to and further away from (i.e. in thez-direction) the focusing system such that the area of high radiationdensity changes position in the flow cell, and v) an optical collectionsystem for collecting and directing at least a portion ofelectromagnetic radiation onto a photodetector, wherein optionally thephotodetector is positioned in optical communication with the flow cellfor receiving incident electromagnetic radiation transmitted through theflow cell and electromagnetic radiation forwarded scattered by theparticle; wherein the photodetector produces an electric signalcharacteristic of the number and/or size of the particles detected;wherein the change in position of the area of high radiation densityallows for characterization of the particles in a larger cross sectionalarea of the flow cell and/or in a larger volume of fluid. In someembodiments, for example, the optical detector array is provided at ascattering angle that is within 5 degrees of zero degrees relative tothe optical axis of the incident beam, and optionally provided at ascattering angle that is within 0.5 degrees of zero degrees relative tothe optical axis of the incident beam.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation to generate an area of high radiation density,such as a focused beam region, within the flow cell; iv) a translator,such as an oscillator, operably connected to the flow cell fortranslating the flow cell laterally (e.g. in a direction orthogonal tothe probe beam axis) across the beam of electromagnetic radiation suchthat the area of high radiation density changes position in the flowcell, and v) an optical collection system for collecting and directingat least a portion of electromagnetic radiation onto a photodetector,wherein optionally the photodetector is positioned in opticalcommunication with the flow cell for receiving incident electromagneticradiation transmitted through the flow cell and electromagneticradiation forwarded scattered by the particle; wherein the photodetectorproduces an electric signal characteristic of the number and/or size ofthe particles detected; wherein the change in position of the area ofhigh radiation density allows for characterization of the particles in alarger cross sectional area of the flow cell and/or in a larger volumeof fluid. In some embodiments, for example, the optical detector arrayis provided at a scattering angle that is within 5 degrees of zerodegrees relative to the optical axis of the incident beam, andoptionally provided at a scattering angle that is within 0.5 degrees ofzero degrees relative to the optical axis of the incident beam, andoptionally for some applications, the photodetector is a differentialdetection system.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation to generate an area of high radiation density,such as a focused beam region within the flow cell; iv) a firsttranslator, such as an oscillator, operably connected to the flow cellfor translating the flow cell closer to and further away from thefocusing system such that the area of high radiation density changesdepth in the flow cell, v) a second translator, such as an oscillator,operably connected to the flow cell for translating the flow celllaterally across the beam of electromagnetic radiation such that thearea of high radiation density changes lateral position in the flowcell, and vi) an optical collection system for collecting and directingat least a portion of electromagnetic radiation onto a photodetector,wherein optionally the photodetector is positioned in opticalcommunication with the flow cell for receiving incident electromagneticradiation transmitted through the flow cell and electromagneticradiation forwarded scattered by the particle; wherein the photodetectorproduces an electric signal characteristic of the number and/or size ofthe particles detected; wherein the first translator and the secondtranslator operate independently and the change in depth and lateralposition of the area of high radiation density allows forcharacterization of the particles in a larger volume of the flow cell.In some embodiments, for example, the optical detector array is providedat a scattering angle that is within 5 degrees of zero degrees relativeto the optical axis of the incident beam, and optionally provided at ascattering angle that is within 0.5 degrees of zero degrees relative tothe optical axis of the incident beam, and optionally for someapplications, the photodetector is a differential detection system.

The translator, such as an oscillator, may translate the flow cell alonga periodic displacement or oscillate at a frequency higher than the timerequired for the particles to pass through the area of high radiationdensity.

Adjusting the profile of the beam at the waist (the narrowest point ofthe propagating beam and therefore the highest in energy density) mayalso be used to increase the cross-sectional area of the flow cell beinganalyzed. For example, by enlarging the beam waist in the lateraldirection (x) a higher cross-sectional area or volume may pass throughthe waist, which typically provides the necessary energy density todetect particles in the fluid. The energy density or laser power willdecrease as the area is enlarged, however by reducing the beam waist inthe vertical direction (y), a high energy density may be maintained.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation into the flow cell, wherein the focusingsystem generates a double waist of the electromagnetic beam in both thex-direction and the y-direction on a transverse plane within the flowcell; wherein the double waist has a greater length in the in thex-direction than in the y-direction; iv) an optical collection systemfor collecting and directing at least a portion of electromagneticradiation onto a photodetector, wherein optionally the photodetector ispositioned in optical communication with the flow cell for receivingincident electromagnetic radiation transmitted through the flow cell andelectromagnetic radiation forwarded scattered by the particle; whereinthe photodetector produces an electric signal characteristic of thenumber and/or size of the particles detected. The double waist has alength in the x-direction greater than or equal to 2 times, 10 times, 20times, 50 times, or optionally, 100 times the length in the y-direction.In some embodiments, for example, the optical detector array is providedat a scattering angle that is within 5 degrees of zero degrees relativeto the optical axis of the incident beam, and optionally provided at ascattering angle that is within 0.5 degrees of zero degrees relative tothe optical axis of the incident beam, and optionally for someapplications, the photodetector is a differential detection system

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation to generate an area of high radiation density,such as a focused beam region, within the flow cell; and iv) an opticalcollection system for collecting and directing at least a portion ofelectromagnetic radiation onto a pixelated photodetector, whereinoptionally the pixelated photodetector is positioned in opticalcommunication with the flow cell for receiving incident electromagneticradiation transmitted through the flow cell and electromagneticradiation forwarded scattered by the particle, wherein each pixel of thepixelated photodetector has an area corresponding to a spatial extent ofthe particle beam interaction signal within the beam; wherein thepixelated photodetector produces an electric signal characteristic ofthe number and/or size of the particles detected. In some embodiments,an area corresponding to a spatial extent of the particle beaminteraction signal within the beam refers to an area that is 75% matchedto the spatial extent of the particle beam interaction signal,optionally for some applications an area that is 90% matched to thespatial extent of the particle beam interaction signal, and optionallyfor some applications an area that is 95% matched to the spatial extentof the particle beam interaction signal. In some embodiments, forexample, the optical detector array is provided at a scattering anglethat is within 5 degrees of zero degrees relative to the optical axis ofthe incident beam, and optionally provided at a scattering angle that iswithin 0.5 degrees of zero degrees relative to the optical axis of theincident beam, and optionally for some applications, the photodetectoris a differential detection system.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation to generate an area of high radiation density,such as a focused beam region, within the flow cell; iv) and an opticalcollection system for collecting and directing at least a portion ofelectromagnetic radiation onto a pixelated photodetector, whereinoptionally the photodetector is positioned in optical communication withthe flow cell for receiving incident electromagnetic radiationtransmitted through the flow cell and electromagnetic radiationforwarded scattered by the particle, wherein the optical collectionsystem recollimates or focuses the beam of electromagnetic radiation;wherein each pixel of the pixelated photodetector has an areacorresponding to a spatial extent of the particle beam interactionsignal within the beam; wherein the photodetector produces an electricsignal characteristic of the number and/or size of the particlesdetected. In some embodiments, for example, the optical detector arrayis provided at a scattering angle that is within 5 degrees of zerodegrees relative to the optical axis of the incident beam, andoptionally provided at a scattering angle that is within 0.5 degrees ofzero degrees relative to the optical axis of the incident beam.

The image of the particle-beam interaction signal is important in theslow axis (long axis) of the beam at the detector. The vertical extentof the signal in the beam is less important for some methods andapplications. The signal will transition across the upper and lowerdetector elements as the particle transits the beam. To maximizesignal-to-noise, the spatial extent of the particle-beam interactionsignal in the slow axis may be predominantly located on a single pair ofdetector elements. Dispersing the particle-beam interaction signalacross multiple pairs of detectors will reduce the signal-to-noise ofthe measurement.

In some embodiments, the pixelated photodetector characterizes particlesbased on one or more horizontal rows of pixels (e.g., distinguishesactual particles from noise), for example, a horizontal row having aheight of 100 pixels, 20 pixels, 10 pixels, 5 pixels, 3 pixels, 2 pixelsor optionally 1 pixels. For differential detection, two or more rows ofhorizontal pixels may be used.

The systems and methods described herein may also be used to determineor estimate the refractive index of a particle, for example, relative tothe refractive index of the fluid media. The system may distinguishbiological particles, such as cells and microbial particles, fromnon-biological particles using a side scatter detector. The refractiveindex of biological particles tends to be relatively similar to thefluid being analyzed, because cells and cell fragments contain a largepercentage of water. Thus, by including a side scatter detector, adetection event that triggers both the primary photodetector and theside scatter detector will correspond to a non-biological particle asthe radiation will be refracted or scattered into the side detector.However, a detection event that triggers the primary detector but doesnot trigger the side scatter will correspond to a biological particle,because the radiation will not be refracted to the extent necessary todirect it towards the side detector.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation into the flow cell; iii) an optical collectionsystem for collecting and directing at least a portion ofelectromagnetic radiation onto a photodetector, wherein optionally thephotodetector is positioned in optical communication with the flow cellfor receiving incident electromagnetic radiation transmitted through theflow cell and electromagnetic radiation forwarded scattered by theparticle; iv) a side scatter detector in optical communication with theflow cell; wherein the photodetector produces an electric signalcharacteristic of the number and/or size of the particles detected;wherein the side scatter detector allows the system to characterize aparticle a biological or non-biological due to the difference ofrefractive index of the fluid and the particle. In some embodiments, forexample, the optical detector array is provided at a scattering anglethat is within 5 degrees of zero degrees relative to the optical axis ofthe incident beam, and optionally provided at a scattering angle that iswithin 0.5 degrees of zero degrees relative to the optical axis of theincident beam.

The systems using differential detection as described herein may alsocharacterize the refractive index of a particle as being higher or lowerthan the refractive index of the fluid. This is important, as metalparticles will typically have a refractive index less than that ofcommon fluids, while non-metals will have a refractive index higher thanthe fluid. Most metals are conductive and conductive materials are moreharmful in many stages of the semiconductor manufacturing process, thus,distinguishing metals help identify more dangerous particles.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a liquid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation into the flow cell; iv) an optical collectionsystem for collecting and directing at least a portion ofelectromagnetic radiation onto a photodetector comprising at least twodetector elements, wherein optionally the photodetector is positioned inoptical communication with the flow cell for receiving incidentelectromagnetic radiation transmitted through the flow cell andelectromagnetic radiation forwarded scattered by the particle; whereineach detector element produces an electric signal characteristic of thenumber and/or size of the particles detected and the photodetectorcharacterizes the particles based on a differential signal generatedfrom each detector element signal; wherein the detector characterizesthe particles as having a lower or higher refractive index than thefluid. The detector may characterize the particle as a metal or anon-metal. In some embodiments, for example, the optical detector arrayis provided at a scattering angle that is within 5 degrees of zerodegrees relative to the optical axis of the incident beam, andoptionally provided at a scattering angle that is within 0.5 degrees ofzero degrees relative to the optical axis of the incident beam.

In an aspect, provided is an system for detecting particles in a fluid,the system comprising: i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation, optionally astructured beam such as a dark beam; iii) a focusing system in opticalcommunication with the optical source for focusing the beam ofelectromagnetic radiation into the flow cell; iv) an optical collectionsystem for collecting and directing at least a portion ofelectromagnetic radiation onto a photodetector, wherein thephotodetector comprises at least two detection elements, whereinoptionally the photodetector is positioned in optical communication withthe flow cell for receiving incident electromagnetic radiationtransmitted through the flow cell and electromagnetic radiationforwarded scattered by the particle; and v) an adjuster operablyconnected to the photodetector or to the focusing system; wherein theadjuster is moves the photodetector or alters the focusing system suchthat the intensity of the electromagnetic beam is distributed in onedirection evenly over each of the detection elements of thephotodetector; wherein the photodetector produces an electric signalcharacteristic of the number and/or size of the particles detected. Thebeam may be distributed vertically, horizontally or both. In someembodiments, for example, the optical detector array is provided at ascattering angle that is within 5 degrees of zero degrees relative tothe optical axis of the incident beam, and optionally provided at ascattering angle that is within 0.5 degrees of zero degrees relative tothe optical axis of the incident beam.

In an aspect, a system for detecting particles in a fluid is provided,the system comprising: (i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, (ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation; (iii) afocusing system in optical communication with the optical source forfocusing the beam of electromagnetic radiation to generate an area ofhigh radiation density within the flow cell; and (iv) an opticalcollection system for collecting and directing at least a portion ofelectromagnetic radiation onto a pixelated photodetector, wherein for atleast a portion of the pixels of the pixelated photodetector each pixelhas an area sufficient to collect the majority of the energy of theparticle-beam interaction signal; (v) wherein the pixelatedphotodetector produces an electric signal characteristic of the numberand/or size of the particles detected.

In an aspect, a system for detecting particles in a fluid is provided,the system comprising: (i) a flow chamber for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, (ii) an optical source in optical communication with the flowchamber for providing the beam of electromagnetic radiation; (iii) afocusing system in optical communication with the optical source forfocusing the beam of electromagnetic radiation to generate an area ofhigh radiation density within the flow chamber; and (iv) an opticalcollection system for collecting and directing at least a portion ofelectromagnetic radiation onto a pixelated photodetector, wherein for atleast a portion of the pixels of the pixelated photodetector each pixelhas an area corresponding to a spatial extent of the particle beaminteraction signal within the beam; wherein the pixelated photodetectorproduces an electric signal characteristic of the number and/or size ofthe particles detected.

In an aspect, a system for detecting particles in a fluid is provided,the system comprising: (i) a flow cell for flowing a fluid containingparticles along a flow direction through a beam of electromagneticradiation, (ii) an optical source in optical communication with the flowcell for providing the beam of electromagnetic radiation; (iii) afocusing system in optical communication with the optical source forfocusing the beam of electromagnetic radiation to generate an area ofhigh radiation density within the flow cell; and (iv) an opticalcollection system for collecting and directing at least a portion ofelectromagnetic radiation onto a pixelated photodetector, wherein theoptical collection system recollimates or focuses the beam ofelectromagnetic radiation; wherein for at least portion of the pixels ofthe pixelated photodetector each pixel has an area corresponding to aspatial extent of the particle beam interaction signal within the beam;wherein the photodetector produces an electric signal characteristic ofthe number and/or size of the particles detected xx

The adjuster may be operably connected to the photodetector andtranslates, moves, rotates or tilts the photodetector. The adjuster maybe a mirror or a lens operably connected to the focusing system andadjusts a path of the beam of electromagnetic radiation. The adjustermay also be configured to provide optical beam power density to the flowcell or to the photodetector, adjust the beam spot size, adjust an areaof high radiation density in the flow cell or any combination thereof.The systems described herein may further comprise an imager, wherein thebeam of electromagnetic radiation may be directed toward the imager andthe imager provides feedback to the adjuster in a closed loop on optimaloptical beam power density, optimal beam spot size, optimal area of highradiation density in the flow cell or any combination thereof.

The beam of electromagnetic radiation may be a Gaussian beam, astructured non-Gaussian beam, a structured dark beam or an anamorphicbeam in a top hat configuration.

System of the invention include optical particle counters, opticalparticle analyzers and optical particle size classifiers.

The photodetector may comprise at least two detector elements andcharacterizes the particles based on a differential signal fromindividual signals from each detector element indicative of theparticles. The described systems may comprise an analyzer for generatingand/or analyzing the differential signal. The focusing system may directthe beam of electromagnetic radiation through the flow cell at leasttwice and the particles in the flow cell interact with a differentportion of the beam on each individual pass through the flow cell. Theanalyzer may analyze the differential signal in the time domain. Thefocusing system may comprise a half wave plate, a quarter wave plate orboth for altering a polarization state of the beam.

The described systems may further comprise a modulator in opticalcommunication with the optical source to modulate the beam ofelectromagnetic radiation, for example, a modulator such as a chopper.The modulator may have a frequency of modulation greater than or equalto 50 kHz, 100 kHz, 200 kHz or optionally, 500 kHz. The photodetectormay have a cooling system to reduce dark current, thereby increasingsignal-to-noise ratio. The described systems may further comprise alock-in amplifier, wherein the lock-in amplifier is bandwidth tuned tothe frequency of the modulator.

The focusing system may comprise one or more diffractive opticalelements. The diffractive optical element may elongate a depth of focusof the beam of electromagnetic radiation thereby generating a longerbeam waist and a larger area of high radiation density within the flowcell.

The focusing system may comprise a varifocal lens for modifying a depthof focus or an area of high radiation density in the flow cell, forexample, an ultra-fast varifocal lens.

The various aspects described herein may also be used in combinationwith one another, and the various possible combinations are specificallydisclosed herein.

Various methods of use of the described systems for the detection ofparticles in a fluid are also specifically disclosed herein.

The invention also provides methods for detecting, counting and/orcharacterizing the size of particles in a fluid using a probe beam ofelectromagnetic radiation, such as a structured beam including a darkbeam. In some embodiments, the methods comprise detection of transmittedand forward scattered light from a flow cell having the fluid with theparticles. In some embodiments, the methods comprise detection oftransmitted and forward scattered light from a flow cell using adifferential detection configuration, such as differential detectionusing a segmented detector comprising one or more pixel pairs, and/orusing a structured beam such as a dark beam. In some embodiments, themethods comprise detection of transmitted and forward scattered lightfrom a flow cell having the fluid with the particles, optionally withadditional off-axis detection of scattered light, for example usingscatter light collection options and scattered light detector. Theinvention also provides methods for detecting, counting and/orcharacterizing the size of particles in a fluid by detection oftransmitted and forward scattered light optionally providing for rapidtranslation (e.g., faster than the average velocity of the particle asit passes through the beam) of the flow cell laterally and/or along thez-axis (e.g., along the beam axis between the light source and detector)to achieve greater sampled volume of fluid per unit time relative tosystems not employing translation. The invention also provides methodsfor detecting, counting and/or characterizing the size of particles in afluid by detection of transmitted and forward scattered light optionallyusing pixelated photodetector(s) having pixel areas corresponding to(e.g., within a factor of 1.5 and optionally a factor of 1.2) thespatial extent of the particle beam interaction signal within the beam,thereby providing an enhancement for detection of more particletransitions leading to an improvement of the sample volume analyzed perunit time. The invention also provides methods for detecting, countingand/or characterizing the size of particles in a fluid by detection oftransmitted and forward scattered light optionally including additionaloff-axis detection of scattered light or fluorescence for distinguishingbetween biological and non-biological particles, for example usingscatter light collection options and scattered light detector.

In an embodiment, a method for detecting, counting and/or characterizingthe size of particles in a fluid comprises the steps: (i) providing aflow of the fluid containing particles, for example in a flow cell; (ii)generating a beam of electromagnetic radiation using an optical source,and optionally one or more beam steering and/or shaping components, suchas a structured beam or dark beam; (iii) passing the beam ofelectromagnetic radiation through the flow cell, for example using abeam steering and/or shaping optical system such as a focusing system,thereby generating electromagnetic radiation transmitted by the flowcell and electromagnetic radiation forward scattered by a particle(s) inthe flow cell; (iv) directing at least a portion of electromagneticradiation transmitted by the flow cell and electromagnetic radiationforward scattered by the particle from the flow cell onto an opticaldetector array, such as a segmented optical detector array comprisingone or more pixel pairs' (v) detecting portion of electromagneticradiation transmitted by the flow cell and electromagnetic radiationforward scattered by the particle, thereby generating one or moresignals, and (vi) analyzing the one or more signal, for example usinghardware or a processor, such as generating and analyzing a differentialsignal, thereby detecting and/or analyzing the particles. In anembodiment of the methods the optical detector array is positioned inoptical communication with the flow cell for receiving incidentelectromagnetic radiation transmitted through the flow cell andelectromagnetic radiation scattered by the particle, for example,wherein the electromagnetic radiation scattered by the particlecomprises forward scattered electromagnetic radiation. In an embodimentof the methods, the incident electromagnetic radiation transmittedthrough the flow cell and the electromagnetic radiation scattered by theparticle undergo constructive and/or destructive optical interference.In an embodiment of the methods the optical detector array is providedat a scattering angle that is within 5 degrees of zero degrees relativeto the optical axis of the incident beam, optionally for someapplications at a scattering angle that is within 1 degree of zerodegrees relative to the optical axis of the incident beam, optionallyfor some applications at a scattering angle that is within 0.5 degree ofzero degrees relative to the optical axis of the incident beam, andoptionally for some applications at a scattering angle that is within0.1 degree of zero degrees relative to the optical axis of the incidentbeam. In an embodiment of the methods, the optical detector array isprovided in optical communication with the flow cell for detecting theinteraction of the particle and the electromagnetic radiation scatteredby the particle with the illuminating wave front.

Without wishing to be bound by any particular theory, there may bediscussion herein of beliefs or understandings of underlying principlesrelating to the devices and methods disclosed herein. It is recognizedthat regardless of the ultimate correctness of any mechanisticexplanation or hypothesis, an embodiment of the invention cannonetheless be operative and useful.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A. and 1B provides schematics of system and method for on-axisparticle measurements by detection of transmitted and forward scatteredlight, for example using a structured beam such as a dark beam, andoptionally a differential detection configuration. FIG. 1A shows asingle pass optical geometry and FIG. 1B shows a dual pass opticalgeometry.

FIG. 2 shows a perspective view of components of a particle measuringsystem illustration an oscillating transducer operably connected to asample cell which is in optical communication with an objective lens.

FIG. 3 provides a schematic showing an example pixelated, differentialdetector configuration.

FIG. 4 provides a schematic showing optical geometry and detectorconfigurations for measurement of refractive index, including examplesignals showing “flipping of the classical particle signal” so as todistinguish between different particle composition.

FIG. 5 provides a schematic showing optical geometry and detectorconfigurations for providing closed loop feedback control ofdifferential detector alignment.

DETAILED DESCRIPTION OF THE INVENTION

In the following description, numerous specific details of the devices,device components and methods of the present invention are set forth inorder to provide a thorough explanation of the precise nature of theinvention. It will be apparent, however, to those of skill in the artthat the invention can be practiced without these specific details.

In general, the terms and phrases used herein have their art-recognizedmeaning, which can be found by reference to standard texts, journalreferences and contexts known to those skilled in the art. The followingdefinitions are provided to clarify their specific use in the context ofthe invention.

“Particles” refers to small objects which are often regarded ascontaminants. A particle can be any material created by the act offriction, for example when two surfaces come into mechanical contact andthere is mechanical movement. Particles can be composed of aggregates ofmaterial, such as dust, dirt, smoke, ash, water, soot, metal, oxides,ceramics, minerals, or any combination of these or other materials orcontaminants. “Particles” may also refer to biological particles, forexample, viruses, spores and microorganisms including bacteria, fungi,archaea, protists, other single cell microorganisms. In someembodiments, for example, biological particles are characterized by asize dimension (e.g., effective diameter) ranging from 0.1-15 μm,optionally for some applications ranging from 0.5-5 μm. A particle mayrefer to a small object which absorbs, emits or scatters light and isthus detectable by an optical particle counter. As used herein,“particle” is intended to be exclusive of the individual atoms ormolecules of a carrier fluid, for example water, air, process liquidchemicals, process gases, etc. In some embodiments, particles may beinitially present on a surface, such as a tools surface in amicrofabrication facility, liberated from the surface and subsequentlyanalyzed in a fluid. Some systems and methods detect particlescomprising aggregates of material having a size dimension, such aseffective diameter, greater than 5 nm, 10 nm, 20 nm, 30 nm, 50 nm, 100nm, 500 nm, 1 μm or greater, or 10 μm or greater. Some embodiments ofthe present invention detect particles having a size dimension, such aseffective diameter, selected from that range of 10 nm to 150 μm,optionally for some applications 10 nm to −10 μm, optionally for someapplications 10 nm to −1 μm, and optionally for some applications 10 nmto −0.5 μm.

The expression “detecting a particle” broadly refers to sensing,identifying the presence of, counting and/or characterizing a particle,such as characterizing a particle with respect to a size dimension, suchas effective diameter. In some embodiments, detecting a particle refersto counting particles. In some embodiments, detecting a particle refersto characterizing and/or measuring a physical characteristic of aparticle, such as effective diameter, cross sectional dimension, shape,size, aerodynamic size, or any combination of these. In someembodiments, detection a particle is carried out in a flowing fluid,such as gas having a volumetric flow rate selected over the range of0.05 CFM to 10 CFM, optionally for some applications 0.1 CFM to 5 CFMand optionally for some applications 0.5 CFM to 2 CFM. In someembodiments, detection a particle is carried out in a flowing fluid,such as liquid having a volumetric flow rate selected over the range of1 to 1000 m L/min.

“Optical Particle Counter” or “particle counter” are usedinterchangeably and refer to a particle detection system that usesoptical detection to detect particles, typically by analyzing particlesin a fluid flow. Optical particle counters include liquid particlecounters and aerosol particle counters, for example, including systemsfor detecting individual single particles in a fluid flow. Opticalparticle counters provide a probe beam of electromagnetic radiation(e.g., a laser) into the analysis area or volume, where the beaminteracts with any particles and then detects the particles based onscattered (forward and/or side scatter), emitted and/or transmittedlight from the flow cell. Detection may focus on electromagneticradiation that is scattered, absorbed, obscured and/or emitted by theparticle(s). Various detectors for optical particle counters are knownin the art, including for example, single detection elements (e.g.,photodiode, photomultiplier tube, etc.), detector arrays includingsegmented detectors, cameras, various detector orientations, etc.Optical particle counter includes condensation particle counters,condensation nuclei counters, split beam differential systems and thelike. When used in the context of a condensation particle counter, theparticle counter portion typically refers to the detection system orcomponents thereof (e.g., source of electromagnetic radiation, optics,filters, optical collection, detector, processor, etc.). In anembodiment, for example, an optical particle counter comprises a sourcefor generating a beam of electromagnetic radiation, beam steering and/orshaping optics for directing and focusing the beam into a region where afluid sample is flowing, for example a liquid or gas flowing through aflow cell. A typical optical particle counter comprises of aphotodetector, such as optical detector array in optical communicationwith said flow cell, and collection optics for collecting and imaginingelectromagnetic radiation which is scattered, transmitted by or emittedby particles which pass through the beam. Particle counters may furthercomprise electronics and/or processors components for readout, signalprocessing and analysis of electrical signals produced by thephotodetector including current to voltage converters, pulse heightanalyzers, and signal filtering and/or amplification electronics. Anoptical particle counter may also comprise a fluid actuation systems,such as a pump, fan or blower, for generating a flow for transporting afluid sample containing particles through the detection region of a flowcell, for example, for generating a flow characterized by a volumetricflow rate. Useful flow rates for samples comprising one or more gasesinclude a flow rate selected over the range of 0.05 CFM to 10 CFM,optionally for some applications 0.1 CFM to 5 CFM and optionally forsome applications 0.5 CFM to 2 CFM. Useful flow rates for samplescomprising one or more liquids include a flow rate selected over therange of 1 to 1000 m L/min.

The expression “interferometric detection of particles” refers tosystems and methods using optical interference to detect one or moreparticles. In some embodiments, coherent beams of electromagneticradiation are superimposed to cause optical interference for sensing,counting and/or determining a size characterization of a particle thatinteracts with at least a portion of the electromagnetic radiation.

“Structured beam detection” refers to systems and methods wherein astructured beam of electromagnetic radiation having a non-Gaussianintensity distribution is passed through a flow cell containing aparticle and is detected using an optical detector array to sense, countand/or characterize the particle.

“Dark beam detection” refers to systems and methods wherein a dark beamof electromagnetic radiation, for example having a spatial intensityprofile having region of attenuated intensity such as a centerlinedecrease in intensity, is passed through a flow cell containing aparticle and is detected using an optical detector array to sense, countand/or characterize the particle.

Detecting and counting small particles (e.g., effective diameter lessthan 100 nm) in clean and ultraclean fluids in a manner that providesstatistically significant data requires high signal-to-noise ratio(abbreviated as S/N or SNR). A high S/N ratio allows nanoparticles to beclearly detected above the noise floor. As used herein “statisticallysignificant data” refers to detection of enough particles per unit timeto be able to accurately assess contamination levels in the fluid. Insome embodiments, high S/N does not relate to sizing accuracy directly.For example, in some optical particle counters the beam waist occupies asmall fraction of the flow cell channel, and therefore, this approachmonitors a subset of the total flow, such that it is possible forparticles to pass through the edge of the beam where irradiance is lessthan the center. If a 50 nm particle passes through the outer edge ofthe beam, it may generate a signal similar to a 10 nm particle passingthrough the center of the beam. Therefore, it is possible form someoptical particle counters to have high S/N and be able to detect 2 nmparticles, while not having very good sizing accuracy. In some of thepresent optical particle counters and methods a goal is to be able tocount enough particles to provide a quantitative, statistically sound,assessment of contamination levels in ultrahigh purity fluids in theshortest period of time. For example, the current state of the artparticle counter may require up to 40 minutes to count enough particlesto provide a statistically appropriate concentration (acceptablerelative standard deviation) measurement when monitoring a state of theart ultrapure water system. By improving and maintaining a high S/Nthrough the present systems and methods, the time interval needed tomeasure this minimum statistically acceptable particle counts can bereduced by 10 x or more. This provides value as it allows a user toidentify deviations from process control limits more quickly.

The expression “high signal-to-noise ratio” refers to a signal-to-noiseratio of an optical particle detection system sufficient for accurateand sensitive detection of particles in a fluid flow, includingparticles characterized by a small physical dimension (e.g., aneffective diameter of less than or equal to 200 nm, optionally for someembodiments less than or equal to 100 nm and optionally for someembodiments less than or equal to 50 nm). In an embodiment, “highsignal-to-noise ratio” refers to a signal-to-noise ratio sufficientlyhigh to sense particles characterized by a small physical dimension,such as particles having an effective diameter as low as 20 nm,optionally for some applications a diameter as low as 10 nm andoptionally for some applications a diameter as low as 1 nm. In anembodiment, “high signal-to-noise ratio” refers to a signal-to-noiseratio sufficiently high to accurately detect and count particles with afalse detection rate of less than or equal to 50 counts/L, for example,for detection of particles having an effective diameter selected overthe range of 1-1000 nm. In an embodiment, “high signal-to-noise ratio”refers to a signal-to-noise ratio sufficiently high to provide a minimumstatistically acceptable particle count in a timeframe at least a factorof 10× less than in a conventional optical particle counter.

The expression “differential detection” refers to techniques and systemsusing the differential signal from forward looking on axis detectorpair(s) for example, at a scattering angle that is within 0.5 degrees ofzero degrees relative to the optical axis of the incident beam,optionally for some applications at a scattering angle that is within0.1 degree of zero degrees relative to the optical axis of the incidentbeam, and optional is at or near zero degrees. A minimum of two pixelscan be used to generate a differential signal (e.g., one upper (or top)and one lower (or bottom)), thereby forming a single pixel pair fordifferential detection. Alternatively, a plurality of pixels may beemployed for each active detector area of a differential detector (e.g.,the top active region and the bottom active region), such as a segmenteddifferential detector comprising one or more pixel pairs, thereby usinga plurality of pixel pairs, for example, wherein one pixel of each pixelpair corresponds to top active detector region and the other pixel ofeach pixel pair corresponds to the bottom active region. The number ofpixel pairs may range, for example, from 1 to 500 pixels and, andoptionally for some applications from 50-100 pixels. In some embodiment,the differential signal is generated by differentially adding signalsfrom pixel pairs corresponding to different active regions of asegmented detector array, such as the top half and the bottom half.Differential detection may be used in the present systems and methods toprovide a reduction of noise and thus enhanced signal-to-noise ratio. Insome embodiments, for example, differential detection is used fordetection of the combination of incident electromagnetic radiationtransmitted through said flow cell and electromagnetic radiation forwardscattered by one or more particle(s) in the fluid flow in the flow cell.In some embodiments, for example, the distribution of light incident hasa power distribution that is balanced between the first and secondactive detection regions (e.g., upper half and lower half) of thedifferential detector, for example, such that the first and secondactive detection regions are characterized by incident radiant powersthat are within 10%, optionally for some applications 5% and optionallyfor some application within 1%. Differential detection includestechniques and systems having closed loop control, for example, based onan evaluation the noise amplitude of the differential signal whenparticles are not present (i.e., in the absence of scattering form theparticle). In some embodiments, a steering mirror is used to adjust theincident beam position on the detector to reduce or minimize noiselevels of the differential signal, which may occur when the beam poweris uniformly split between the first and second active detector elements(e.g., upper and lower elements of the detector). Close loop control canalso be achieved by translating the detector position and rotating thedetector to align the beam and detector axes so as to reduce or minimizenoise levels of the differential signal.

“Structured Beam” refers to a coherent beam of electromagnetic radiation(e.g., a laser) having a non-Gaussian spatial intensity distribution.Structured beams include beams characterized by an attenuated region,such as a dark beam, beams with a line focus with a dark linesingularity, beams characterized by two or more discrete intensitylobes, etc. In an embodiment, a structured beam corresponds to atransverse mode, such as a TEM01. Structured beams include focused,synthesized, laser beams. Structured beams and dark beams may begenerated by techniques known in the art including using an opticalmask, modification of a laser cavity, combining multiple beams, spatialand/or polarization filters and other manipulations such as in aninterferometric or polarization modification scheme.

“Beam propagation axis” refers to an axis parallel to the direction oftravel of a beam of electromagnetic radiation.

“Optical communication” refers to components which are arranged in amanner that allows light to transfer between the components. Opticalcommunication includes configurations where two elements are directly inoptical communication wherein light travels directly between theelements and configurations where two elements are indirectly in opticalcommunication wherein light travels between the elements via one or moreadditional optical elements, such as lenses, mirror, windows, filters,etc.

“Optical axis” refers to a direction along which electromagneticradiation propagates through a system.

“Optical detector array” refers to an optical detector for spatiallyresolving input signals (e.g., electromagnetic radiation) in twodimensions across an active area of the detector. An optical detectorarray may generate an image, for example an image corresponding to anintensity pattern on the active area of the detector. In an embodiment,an optical detector array comprises an array of individual detectorelements, also referred herein as pixels; for example: a two-dimensionalarray of photodetectors, a charge-coupled device (CCD) detector, acomplementary metal-oxide-semiconductor (CMOS) detector, ametal-oxide-semiconductor (MOS) detector, an active pixel sensor, amicrochannel plate detector, or a two-dimensional array of photodiodes.

“Optical source” refers to a device or device component for deliveringelectromagnetic radiation to a sample. The term is not limited tovisible radiation, such as by a visible light beam, but is used in abroad sense to include any electromagnetic radiation also inclusive ofvisible radiation, ultraviolet radiation and/or infrared radiation. Theoptical source may be embodied as a laser or laser array, such as adiode laser, diode laser array, diode laser pumped solid state laser,LED, LED array, gas phase laser, solid state laser, to name a fewexamples. In some embodiments, an optical source is for generating oneor more coherent beams of electromagnetic radiation, for example, togenerate a probe beam in an optical particle counter. In an embodiment,an optical source may include one or more components, such as a beamshaping system, phase mask, beam combiner, polarization controller, waveplate, or other component for generating a structured beam, such as adark beam, for providing a probe beam in an optical particle counter.

The term “electromagnetic radiation” and “light” are used synonymouslyin the present description and refer to waves of electric and magneticfields. Electromagnetic radiation useful for the methods of the presentinvention include, but is not limited to ultraviolet light, visiblelight, infrared light, or any combination of these having wavelengthsbetween about 100 nanometers to about 15 microns.

A “high aspect ratio” beam refers to an optical beam, such as a laserbeam including structured beam or dark beam, having an aspect ratioselected from the range of 10:1 to 200:1.

The present systems and methods integrate active and/or passivecomponents for enhancing optical particle detection on-axis particlemeasurements by detection of transmitted and forward scattered light,for example using a structured beam such as a dark beam, and optionallya differential detection configuration to provide important performancebenefits including (i) providing high signal-to-noise ratios andincreased sensitivity for detection and size characterization of smallparticles (e.g., effective lateral dimensions (e.g., diameter) of 10microns, or optionally 1 microns or optionally 500 nanometers), (ii)increasing the amount of sample fluid analyzed as a function of timeand/or (iii) suppressing false positive indications.

FIG. 1 provides a schematic of a system for detection of particles viaon-axis particle measurement by detection of transmitted and forwardscattered light, for example using a structured beam such as a dark beamand a differential detection configuration. A shown in FIG. 1, particledetection system (200) includes a flow cell (210) for transporting flowof fluid (150) including particles (schematically depicted as circleswithin flow cell (210), such as a gas or liquid flow having particles.Optical source (220), such as a laser source, generates electromagneticradiation that is provided to beam steering and shaping system (221) forgenerating a probe beam (222), such as a structured beam including adark beam, that is provided to flow cell (210). The probe beam passesthrough flow cell (210) and is detected via an on-axis optical detectorarray (240), such as segmented 1D or 2D optical detectors (240A and240B) comprising one or more pixel pairs, which is in operationcommunication with processor (101) to provide an output signal(s) toprocessor (101). Optical detector array (240) and/or processor (101) maybe provide differential detection, for example in a configurationwherein individual segmented detector regions are each positioned overdifferent intensity lobes of a structured beam, such as a dark beam.

Processor (101) receives and analyzes the output signals from theoptical detector array (240), for example via generation and analysis ofa differential signal combining (e.g., differentially adding,subtraction, etc.) of signals from segmented 1D or 2D optical detectors(240A and 240B) to provide detection of the particles, such as bycounting and/or size characterization. In some embodiments, one or moretranslators (205), such as an oscillator, displacer, piezoelectricelement, etc., is operationally coupled to flow cell (210) to providefor rapid translation (e.g., at an average translation velocity at least2 times faster than the average velocity of the particle passing throughthe beam) of the flow cell laterally (e.g., in a direction orthogonal tothe axis of the incident probe beam) and/or in the z-axis (e.g., alongthe beam axis between the optical source and the detector and/or alongthe axis of the incident probe beam) to achieve greater sampled volumeof fluid per unit time. In some embodiments, for example, said opticaldetector array (240) is provided at a scattering angle that is within 5degrees of zero degrees relative to the optical axis of the incidentbeam, and optionally provided at a scattering angle that is within 0.5degrees of zero degrees relative to the optical axis of the incidentbeam and optionally provided at a scattering angle that is within 0.1degrees of zero degrees relative to the optical axis of the incidentbeam.

Also shown in FIG. 1A is an optional side scatter photodetector (268)and side scatter collection optics (267) which are positioned off-axisrelative to the beam propagation axis of the probe beam (222) and thedetector axis of optical detector array (240). Side scatter collectionoptics (267), such as one or more lenses and/or mirrors, is positionedto receive off-axis scattered light resulting from interaction with aparticle in flow cell (210) and the probe beam. Side scatter collectionoptics (267) directs, and optionally images, at least a portion ofcollected scattered light on to side scatter photodetector (268) whichis in operation communication to provide an output signal(s) toprocessor (101) for analysis to detect and/or characterize theparticle(s). Embodiments incorporating the combination of on-axisdifferential detection and off-axis side scatter detection areparticularly useful for characterizing particles as biological ornon-biological particles. In some embodiments, for example, processors(101) compares signals from on-axis optical detector array (240) andside scatter photodetector (268), so as to determine if a particle is abiological particle or a non-biological particle. In some embodiments,for example, a small output signal from side scatter photodetector (268)or lack of a measurable signal from side scatter photodetector (268),accompanying a measurable signal from on-axis optical detector array(240) is indicative of a biological particle, such as a microbialparticle.

FIG. 1B provide a schematic of an alternative system for detection ofparticles on-axis particle measurements by detection of transmitted andforward scattered light, for example using a structured beam such as adark beam, and a differential detection configuration, wherein theoptical geometry is set up to provide a dual pass optical geometry. Asshown in FIG. 1B, the system (200) includes optical source (220), beamsteering and shaping system (221), flow cell (210), optical detectorarray (240) comprising a paired detector array and translators (205). Inaddition, beam splitter (265) and mirror (275) are included to provide adual pass optical geometry. Optionally, the beam steering and shapingsystem (221) provides for a high aspect ratio beam, such as a beamcharacterized by an aspect ratio selected from the range of 10:1 to200:1, provided to the flow cell (210) and optical detector array (240)is configured as a paired detector array including paired detectorarrays (240A and 240B, expanded out from their position within detector(240) and schematically illustrated for clarity separately below theparticle detection schematic next to an example signal corresponding toa particle detection event). Optical detector array (240) may beconfigured to provide differential detection, optionally wherein paireddetector arrays (240A and 240B) are positioned over the intensity lobesof a structured beam such as a dark beam. In some embodiments, one ormore translators (205), such as an oscillator, displacer, piezoelectricelement, etc., are operationally coupled to flow cell (210) to providefor rapid translation (e.g., at an average translation velocity 2 timesfaster than the average velocity of the particle passing through thebeam) of the flow cell laterally and/or in the z-axis (e.g., along thebeam axis between the optical source and the detector and/or along theaxis of the incident probe beam) to achieve greater sampled volume offluid per unit time. In some embodiments, for example, said opticaldetector array is provided at a scattering angle that is within 5degrees of zero degrees relative to the optical axis of the incidentbeam, and optionally provided at a scattering angle that is within 0.5degrees of zero degrees relative to the optical axis of the incidentbeam.

FIG. 1B also shows a representative signal of the optical detector array(240) showing a signals from individual paired detector arrays (240A and240B) as a function time (or particle trajectory through the beam) for aparticle passing through the beam in the flow cell, wherein the solidline is the signal from detector array 240A and the dotted line is thesignal from detector array 240B. As shown in FIG. 1B, the signals fromindividual paired detector arrays (240A and 240B) are each characterizedby a minimum value and a maximum value and are inverted with respect toeach other. Signals from individual paired detector arrays (240A and240B) may be combined, for example via differential addition,subtraction, multiplication, etc., to provide a signal, such as adifferential signal, that can be analyzed to provide accurateinformation as to the size, optical properties (e.g., refractive index)and composition of the particle.

The depicted optical geometry allows for constructive and destructiveinterference of the beam which aids in sensitivity, for example,involving the combination of light transmitted from the flow cell andforward light scattered from the particle in flow cell. The use of dualpass optical geometry and differential detection aids in sensitivity andaccuracy for detection of small particles (e.g., have effectivedimensions less than 100 nm, optionally less than 50 nm and optionalless than 20 nm)). The use of a high aspect ratio beam increases thesample volume for which particles may be detected, which increase theamount of sample that can be monitored per unit time.

FIG. 2 shows a perspective view of components of a particle measuringsystem including an oscillating transducer operably connected to asample cell in optical communication with an objective lens. As show inFIG. 2, objective lens (300) directs and focuses an optical beam (320),such as a structured beam, from an optical source (220) onto flow cell(210) which is operationally coupled to a translator comprising anoscillating transducer (310) for translating the flow cell (210) alonglateral direction (330), which is optionally orthogonal to z-axis (331)corresponding to the propagation axis of the incident beam, such thatthe area of high radiation density changes position in the flow cell,thereby allowing for an increase in volume of analyzed fluid per unittime. In some embodiments, transducer (310) provides for translation ofthe flow cell (210) along the z-axis (331), such that the area of highradiation density changes position in the flow cell, thereby allowingfor an increase in volume of analyzed fluid per unit time. Theelectromagnetic radiation transmitted by the flow cell andelectromagnetic radiation forward scattered by particles in the flowcell are detected by optical detector array (240), such as segmented 1Dor 2D optical detectors (240A and 240B), for example using an on axisdifferential detection system.

FIG. 3 provides a schematic showing an example imaged detector using apixelated, differential detector configuration. Section 1 of FIG. 3,provides a schematic of a beam imaged on a pixelated detector having topactive region (“the top half”) and bottom active region (“the bottomhalf”), wherein the energy of the beam is evenly distributed (50% ineach) between top and bottom detector halves to within ±1% to 5%. Thebeam energy is balanced across the two active regions socorrelated-laser noise is at least partially cancelled usingdifferential detection. In addition, sizing matching of the pixelrelative to the signal (i.e., the spatial extent of signal) optimizessignal-to-noise ratio, for example to provide a high signal-to-noiseratio.

Section 2 of FIG. 3, provides a schematic of a beam imaged on apixelated detector, corresponding to conditions wherein a particleenters the beam in the flow cell, for example entering the beam from thebottom. As shown in Section 2 of FIG. 3, a bright fringe is observed onone, or a subset of pixels, for the top half of the pixelated detectorand dark fringe is observed on one, or a subset of pixels, for thebottom half of the pixelated detector. Section 3 of FIG. 3, provides aschematic of a beam imaged on a pixelated detector, corresponding toconditions wherein a particle is translating through the top of the beamwaist in the flow cell. As shown in Section 3 of FIG. 3, a dark fringeis observed on one, or a subset of pixels, for the top half of thepixelated detector and bright fringe is observed on one, or a subset ofpixels, for the bottom half of the pixelated detector. In thisconfiguration, the differential signal is driven by fractionalfluctuation of power at detector. As pixel size increases, fractionalfluctuation decreases when a particle intersects with the beam. If thepixel gets too small, the power on the detector goes down and signalamplitude goes down even if the fractional fluctuation stays the same.Accordingly, there is an optimum in the middle reflecting this tradeoff.

FIG. 4 provides a schematic showing example signals as a function oftime achieved using differential detection of a particle including theDifferential Signal (500), Top Signal (510) from the pixel or subset ofpixels from the top half of the differential detector and Bottom Signal(520) from the pixel or subset of pixels the bottom half of thedifferential detector. As illustrated in FIG. 4, the Differential Signal(500) is generated by differentially adding the Bottom Signal (520) andthe Top Signal (510). By deriving the Differential Signal (500) from TopSignal (510) and Bottom Signal (520) in this manner, a simultaneousreduction in noise is realized via correlated differential noisecancellation and, thereby enhances overall signal-to-noise ratio.

The Differential Signal (500) may be analyzed to provide information asto the effective size dimension(s) and optical properties (e.g.refractive index) of the particle and to distinguish between differentparticle optical properties such as refractive index and, thus, provideinformation on particle composition. To illustrate this concept,detection of particles having different refractive indexes andcompositions are compared—(i) polystyrene latex (PSL) vs (ii) goldnanoparticle. PSL particles have a refractive index of 1.59 which isgreater than water's refractive index of 1.33, therefore, upon enteringthe beam (e.g., circumstances corresponding to Section 2 of FIG. 4) abright fringe is observed on the top half of the differential detectorand a dark fringe is observed on the bottom half of the differentialdetector; and upon translating through the beam waits (e.g.,circumstances corresponding to corresponding to Section 3 of FIG. 4) adark fringe is observed on the top half of the differential detector anda bright fringe is observed on the bottom half of the differentialdetector. If on the other hand, gold nanoparticles having a refractiveindex less than the refractive water at the wavelength of the probe beamare analyzed using some embodiments of the present differentialdetection methods, an inverted signal relative to PSL particles isobserved, wherein upon entering the beam (e.g., circumstancescorresponding to Section 2 of FIG. 4) a dark fringe is observed on thetop half of the differential detector and a bright fringe is observed onthe bottom half of the differential detector; and upon translatingthrough the beam waits (e.g., circumstances corresponding tocorresponding to Section 3 of FIG. 4) a bright fringe is observed on thetop half of the differential detector and a dark fringe is observed onthe bottom half of the differential detector. In this manner, thesequence and position of bright and dark fringes as observed in thedifferential signal may be used to characterize the refractive index(and composition) of the particles analyzed by the present methods.

1D and 2D detectors, including segmented detectors, are useful fordifferential detection in certain embodiments. With 1D segmenteddetectors two options are useful for some applications: (i) orient thedetector segments vertically or parallel to particle transit through thebeam such that two adjacent pixels can be used as a single pair of upperand lower pixels; or, mount two 1D detectors at 90 degrees to the beamand use a knife edge prism to send the top half of the beam to one 1Ddetector and the lower half of the beam to a second 1D detector. Thenumber of pixel pairs may range, for example, from 1 to 500 pixels and,and optionally for some applications from 50-100 pixels. Pixel widthsselected from the range of 10 to 500 microns are useful in certainembodiments and optionally for some applications selected from the rangeof 50-100 microns.

FIG. 5 provides a schematic showing an optical geometry and detectorconfiguration for providing closed loop feedback control of differentialdetector alignment, for example, to balance beam energy across the twoactive regions of the detector (e.g., the top half and bottom half) solaser noise is at least partially cancelled using differentialdetection. Use of closed loop feedback control in certain embodiments,is useful for correcting alignment drift and/or addressing outsideacoustical or vibration interference. As depicted in FIG. 5, opticalsource (600), such as a laser, provides optical beam (605 a), forexample a structured beam, which is passed through flow cell (620) viasteering and/or focusing optics (610 a). The optical beam (605 a)interacts with particles in a fluid flowing through flow cell (620),thereby generating transmitted electromagnetic radiation and forwardscattered electromagnetic radiation (together 605 b), which is collectedvia collection optics (610 b). Transmitted electromagnetic radiation andforward scattered electromagnetic radiation (605 b) is directed on tomirror (630), optionally steering mirror, which directs at least aportion of transmitted electromagnetic radiation and forward scatteredelectromagnetic radiation (605 b) on to differential detector (640) forexample, a segmented detector having a first active region and a secondactive region (e.g., top half and bottom half). Positioner (650) is inoperationally coupled to differential detector (640) so as to adjust theposition of the differential detector (640), such as to move thedetector laterally and/or vertically or to rotate the detector. In anembodiment, processor (660) is in operational communication with mirror(630) and/or positioner (650) so as to control the relative alignment oftransmitted electromagnetic radiation and forward scatteredelectromagnetic radiation (605 b) on first and second active regions ofdifferential detector (640) such as top half and bottom half ofdifferential detector (640). In an embodiment, processor (660) receivesand analyzes signals corresponding to the first active region and thesecond active region (e.g., top half and bottom half) and determines thedifferential signal.

In order to minimize noise and maximize signal, the power transmittedelectromagnetic radiation and forward scattered electromagneticradiation (605 b) of may be balanced between the first active region andthe second active region (e.g., top half and bottom half) differentialdetector (640). In some embodiments, this is accomplished with a closedloop system wherein the processor analyzes the differential signal whenparticles are not present and minimizes the noise amplitude of thedifferential signal via control of positioner (650) and mirror (630). Insome embodiments, for example, mirror (630) is used to adjust the beamposition on the detector (640) to minimize noise levels of thedifferential signal. This condition occurs when the beam power is mostuniformly split between the upper and lower elements of the differentialdetector. Minimize noise levels of the differential signal can also beachieved, for example, via translating the detector position androtating the detector (640) to align the beam and detector axes usingpositioner (650).

The invention can be further understood by the following non-limitingexamples.

Example 1—Particle Measurement Using Structured Beams and/orDifferential Detection

This example describes optical geometries, detector configurations andsignal analysis techniques allowing for enhancements for particledetection and size characterization corresponding to specificembodiments which are intended to exemplify certain specific features ofthe invention.

Scanning Modulated Focus:

In some embodiments, for example, the system is designed to create aregion of high laser beam optical power density at the point ofmeasurement. In conventional systems, this illumination area istypically constrained by the focus angle of the objective lenses andlimits cross sectional area within the sample cell within which thesmallest particles can be identified and characterized. Use of a highspeed mechanical oscillator, such as a piezoelectric or similar device,can be used to physically move or translate the sample cell closer toand farther away from the objective lens. This mechanical translationmoves the point of highest optical density within the sample cell. Whendone at sufficiently high frequency and faster than the particle transittime of the laser beam, allows a larger cross-sectional area of thesample cell to be characterized for particulates. This approach resultsin an increased sample volume of fluid per unit of time withoutrequiring increases in laser power.

Scanning Modulation Cross Axis:

In some embodiments, for example, the system is designed to create aregion of high laser beam optical power density at the point ofmeasurement. In conventional systems, this area is typically constrainedby the focus angle of the objective lenses and limits cross sectionalarea within the sample cell within which the smallest particles can beidentified and characterized. Use of a high speed mechanical oscillatorsuch as a piezoelectric or similar device can be used to physically moveor translate the sample across the laser beam. This mechanicaltranslation moves the point of highest optical density laterally withinthe sample cell. When done at sufficiently high frequency and fasterthan the particle transit time of the laser beam, allows a largercross-sectional area of the sample cell to be characterized forparticulates. This essentially results in an increased sample volume offluid per unit of time without requiring increases in laser power.

Two-Dimensional Scanning Modulation:

In some embodiments, for example, the scanning modulated focus andscanning modulation cross-axis can be used individually or incombination for increased cumulative effect.

Imaging Sample Volume on Detector:

In some embodiments, for example, this feature derives from therelationship that the signal-to-noise ratio (SNR or S/R) will bemaximized when at least a portion of the pixels of the photodetectoreach has a width sufficient to collect the majority of the energy of theparticle-beam interaction signal. The image of the particle-beaminteraction signal is important in the slow axis (long axis) of the beamat the detector. The vertical extent of the signal in the beam is lessimportant. The signal will transition across the upper and lowerdetector elements as the particle transits the beam. To maximizesignal-to-noise, the spatial extent of the particle-beam interactionsignal in the slow axis may be predominantly located on a single pair ofdetector elements. Dispersing the particle-beam interaction signalacross multiple pairs of detectors will reduce the signal-to-noise ofthe measurement. Considering that the sample volume may be illuminatedwith a high aspect ratio beam—with orthogonal beam waists at thelocation of the sample volume—an image of this location created alongthe beam axis will be similarly shaped and should have sufficientmagnification to distribute the image over an appropriate number ofpixel detector elements.

An alternative detector geometry may include the following features.After the focused beam passes through the sample volume, where thehorizontal and vertical beam waists are in the same transverse plane inthe sample volume, the beam is collected by downstream optics andre-collimated. In this region of the beam the signal of particle eventsoccurring at or near the beam waists is distributed over the transversespan of the collimated beam. To optimize the signal-to-noise ratio, thebeam is brought to a focus onto a pixelated detector, where pixel sizeis the same or smaller than RMS spot size of the focusing optics. Inthis way spatial discrimination of particle events throughout the samplevolume is realized. A minimal detector area (area of a pixel) isutilized to capture a particle event.

Fractional Attenuation Imaging:

In some embodiments, for example, the wide (horizontal) and narrow(vertical) image of the sample volume beam cross-section, being createdby focusing the beam after it passes through the sample volume onto apixelated detector plays a role in the present systems and methods. Thisimage at the detector may be a magnified reproduction of the beamprofile at the sample volume. As a particle transits the beam at or nearthe waist, its image at the detector will be a vertical trajectoryacross the image footprint. In the vein of maximizing thesignal-to-noise ratio as described above, a horizontal slice (possiblysingle-pixel width vertically) of the image footprint may be the meansto achieve an optimal signal-to-noise ratio.

High Aspect-Ratio Beam:

In some embodiments, for example, as the beam is presented to the samplefluid flow it will be shaped and focused by appropriate opticalelements. It will be brought to a tight focus in the direction of fluidflow. If the fluid flows along the y-axis, the beam will be tightlyfocused in the y-direction; the point of tightest focus identifies thelocation of the y-axis beam waist. With the z-axis being along the axisof the beam, then along the x-axis the beam will be much wider than itis in the y-direction, but the beam will also need to be at its minimumx-axis width in the same transverse xy-plane as the y-axis waist. Thebeam shaping optics will be arranged so that the locations of the x-axisand y-axis waists occur in the same xy-plane. This being the case, boththe x- and y-axis beam profiles will converge to their respective waistsin the same xy-plane and will diverge from there in the propagationdirection. This constitutes the high aspect-ratio beam. The co-locationof the waists in the same xy-plane is necessary for downstream imagingconsiderations.

Differential Signal:

In some embodiments, for example, using the differential signal from theforward looking, on axis detector pair(s) at a scattering angle of zerodegrees results in significant noise reduction.

Microbial Detection:

In some embodiments, for example, use of a structured beam or dark beamwith on-axis differential detection is very effective at detectingmicro-organisms in water even though they have a very small index ofrefraction contrast. Micro-organisms are often not typically observedwith conventional light scattering particle counters due to the lowindex of refraction contrast (they contain mostly water). A particlecounter of some embodiments uses a dark beam with on-axis differentialdetection combined with side scatter detection. If both detectors createan appropriately sized signal, the particle is not a micro-organism. Ifon-axis differential detector provides a large signal and the sidescatter provides no response, it is a micro-organism.

RI Differences Between Particle and Media:

In some embodiments, for example, the shape of the differential signalvs. time depends on refractive index of the particle relative to themedia. The differential detection signal is either an increasing signalabove zero followed by a decreasing signal below zero (bump followed bydip) or the opposite with the signal decreasing below zero followed byan increasing signal above zero (dip followed by bump). This signalchanges based on the direction of flow and on whether the top detectoris subtracted from the lower or vice versa. As an example: for a givenconfiguration, if the refractive index of the particle is greater thanthe refractive index of the media the detection signal will be a bumpfollowed by a dip while if the refractive index of the particle is lessthan the media, the signal will be a dip followed by a bump. The typesof materials with a refractive index less than water or fluid chemicalsare gases and many metals. Material with an index of refraction lessthan gases include certain metals (in the visible portion of thespectrum).

Closed Loop Focus System:

In some embodiments, for example, adjusting optical elements in allthree axes provides useful and/or optimal beam power density and/or beamsize. Diverting a small portion of the beam which is going to the photodetector to an imager and using the size, shape, and or power density toprovide conditions for useful and/or optimal particle detection. Thelaser beam may be precisely balanced evenly/equally across the upper andlower elements of the differential detector, in order for the lasernoise to cancel out in the differential signal. In other words, thedetected power may beneficially and/or optimally be the same for theupper and lower detector. This can be accomplished in some embodimentsby translating the detector vertically (also a tilt function for 2Darray to align with high aspect ratio beam) or by steering the laserbeam with a mirror or lens to maximize noise cancelation. In someembodiments, a closed loop system is implemented where the detector andlaser are automatically aligned in an arrangement that minimizesbackground noise by optimizing the “balance” of laser power across thedifferential detector elements. Additionally, a closed loop focus systemmay be used to adjust the 5-axis optical translation stages to provideoptimal beam power density and/or beam spot size. For example, a smallfraction of the beam can be redirected to an imager and the imageobtained can be used to determine the adjustments required to obtainoptimal beam size, shape, and power density.

When the operating temperature of detectors is lowered, a lowering ofthe detectors Dark Current follows. This lowering of Dark Currentimproves the detector's SNR (Signal-to-noise Ratio) and lowers thedetector's NEP (Noise Equivalent Power) making it more sensitive tolower incident light levels. There is a theoretical limit to thisresponse in thermal change and designing a thermal system to cool to thenecessary levels with the required stability may be employed.

Chopper Use:

In some embodiments, for example, further improvement in the extractionof low-level optical signals buried in the noise floor can be realizedby considering the problems inherent in DC techniques and how these canbe reduced through the use of AC methods and a modulated light, orchopper approach. In some embodiments, the chopper is used to turn onand turn off the detector's incident light, changing it from a DCillumination to AC illumination, typically in the KHz range. In someembodiments, using a lock-in amplifier, narrowly bandwidth tuned to thechopper frequency creates synchronous signal detection. Most physicalsystems including the electronic optical detection and amplificationinvolved in particle detection have increased noise as the frequencyapproaches DC. For example, typical Op-amps used in particle detectionhave 1/f noise. By moving the detection measurement away from lowfrequency, or DC noise sources to measurements at the AC chopperfrequency, higher signal-to-noise ratio and detection of much weakersignals typically associated with smaller particles, can be achieved.

Signal Processing Techniques in Particle Signal Detection:

In some embodiments, the path and speed that various particles ofdifferent size and material types take through the impinging lightsource in the time domain are useful considerations for the design andsignal analysis in the present systems and methods. For example, thesampling of these discrete and complex time domain signals can then betransformed into the frequency domain through the Fouriertransformation, or FFT (Fast Fourier Transformation). The decompositionof the complex periodic signal trace of particles provides an equationof the frequency domain signal made up of a set sinusoids with differentamplitudes, frequencies and phase. In some embodiments, a collection, orlibrary of equations of decomposed particle signals collected duringdevelopment can be cataloged and used when the magnitude of the inherentnoise of the system is very close to the magnitude of the particlesbeing witnessed, or low signal-to-noise ratio conditions. In someembodiments, the collected particle signals have inherent structure,unique to the design of the invention, that one can use signalprocessing techniques to uncover the expected signal from theunidentifiable disordered signal representing the contaminating systemnoise.

In some embodiments, the equation models, or filters, are what isbelieved to be the structure of possible signals of interest. In someembodiments, when attempting to identify the structure in an incomingsignal, the mathematical filters are used and imposed on the arrivingsignal. In some embodiments, various techniques of signal processing canbe used including convolving the arriving transformed signal with thecatalog of filters (assuming the arriving and modeled signals are lineartime invariant). In some embodiments, correlation between the arrivingsignal and the catalog of filters can be used. Resent developments inthe Convolutional Sparse Modeling technique can be used together withthresholding, relaxation or approximation. In some embodiments, mutualCoherence can be used to break up the arriving signal into smallerpatches of variable delay (phase) and width (frequency) and testedagainst the catalog of models, with the assumption that the arrivingsignals are piece wise constant signals.

STATEMENTS REGARDING INCORPORATION BY REFERENCE AND VARIATIONS

All references throughout this application, for example patent documentsincluding issued or granted patents or equivalents; patent applicationpublications; and non-patent literature documents or other sourcematerial; are hereby incorporated by reference herein in theirentireties, as though individually incorporated by reference, to theextent each reference is at least partially not inconsistent with thedisclosure in this application (for example, a reference that ispartially inconsistent is incorporated by reference except for thepartially inconsistent portion of the reference).

The following patents and patent applications relate to interferometricparticle detection using a structured beam and are incorporated byreference in their entireties: U.S. Pat. No. 7,746,469; US Publication20170176312; and PCT publication WO 2019/082186.

The terms and expressions which have been employed herein are used asterms of description and not of limitation, and there is no intention inthe use of such terms and expressions of excluding any equivalents ofthe features shown and described or portions thereof, but it isrecognized that various modifications are possible within the scope ofthe invention claimed. Thus, it should be understood that although thepresent invention has been specifically disclosed by preferredembodiments, exemplary embodiments and optional features, modificationand variation of the concepts herein disclosed may be resorted to bythose skilled in the art, and that such modifications and variations areconsidered to be within the scope of this invention as defined by theappended claims. The specific embodiments provided herein are examplesof useful embodiments of the present invention and it will be apparentto one skilled in the art that the present invention may be carried outusing a large number of variations of the devices, device components,methods steps set forth in the present description. As will be obviousto one of skill in the art, methods and devices useful for the presentmethods can include a large number of optional composition andprocessing elements and steps.

As used herein and in the appended claims, the singular forms “a”, “an”,and “the” include plural reference unless the context clearly dictatesotherwise. Thus, for example, reference to “a cell” includes a pluralityof such cells and equivalents thereof known to those skilled in the art.As well, the terms “a” (or “an”), “one or more” and “at least one” canbe used interchangeably herein. It is also to be noted that the terms“comprising”, “including”, and “having” can be used interchangeably. Theexpression “of any of claims XX-YY” (wherein XX and YY refer to claimnumbers) is intended to provide a multiple dependent claim in thealternative form, and in some embodiments is interchangeable with theexpression “as in any one of claims XX-YY.”

When a group of substituents is disclosed herein, it is understood thatall individual members of that group and all subgroups, are disclosedseparately. When a Markush group or other grouping is used herein, allindividual members of the group and all combinations andsub-combinations possible of the group are intended to be individuallyincluded in the disclosure.

Every device, system, formulation, combination of components, or methoddescribed or exemplified herein can be used to practice the invention,unless otherwise stated.

Whenever a range is given in the specification, for example, atemperature range, a time range, or a composition or concentrationrange, all intermediate ranges and subranges, as well as all individualvalues included in the ranges given are intended to be included in thedisclosure. It will be understood that any subranges or individualvalues in a range or subrange that are included in the descriptionherein can be excluded from the claims herein.

All patents and publications mentioned in the specification areindicative of the levels of skill of those skilled in the art to whichthe invention pertains. References cited herein are incorporated byreference herein in their entirety to indicate the state of the art asof their publication or filing date and it is intended that thisinformation can be employed herein, if needed, to exclude specificembodiments that are in the prior art. For example, when composition ofmatter are claimed, it should be understood that compounds known andavailable in the art prior to Applicant's invention, including compoundsfor which an enabling disclosure is provided in the references citedherein, are not intended to be included in the composition of matterclaims herein.

As used herein, “comprising” is synonymous with “including,”“containing,” or “characterized by,” and is inclusive or open-ended anddoes not exclude additional, unrecited elements or method steps. As usedherein, “consisting of” excludes any element, step, or ingredient notspecified in the claim element. As used herein, “consisting essentiallyof” does not exclude materials or steps that do not materially affectthe basic and novel characteristics of the claim. In each instanceherein any of the terms “comprising”, “consisting essentially of” and“consisting of” may be replaced with either of the other two terms. Theinvention illustratively described herein suitably may be practiced inthe absence of any element or elements, limitation or limitations whichis not specifically disclosed herein.

One of ordinary skill in the art will appreciate that startingmaterials, biological materials, reagents, synthetic methods,purification methods, analytical methods, assay methods, and biologicalmethods other than those specifically exemplified can be employed in thepractice of the invention without resort to undue experimentation. Allart-known functional equivalents, of any such materials and methods areintended to be included in this invention. The terms and expressionswhich have been employed are used as terms of description and not oflimitation, and there is no intention that in the use of such terms andexpressions of excluding any equivalents of the features shown anddescribed or portions thereof, but it is recognized that variousmodifications are possible within the scope of the invention claimed.Thus, it should be understood that although the present invention hasbeen specifically disclosed by preferred embodiments and optionalfeatures, modification and variation of the concepts herein disclosedmay be resorted to by those skilled in the art, and that suchmodifications and variations are considered to be within the scope ofthis invention as defined by the appended claims.

1-11. (canceled)
 12. A system for detecting particles in a fluid, the system comprising: a flow cell for flowing a fluid containing particles along a flow direction through a beam of electromagnetic radiation, an optical source in optical communication with the flow cell for providing the beam of electromagnetic radiation; a focusing system in optical communication with the optical source for focusing said beam of electromagnetic radiation to generate an area of high radiation density within said flow cell; and an optical collection system for collecting and directing at least a portion of electromagnetic radiation onto a pixelated photodetector, wherein for at least a portion of the pixels of said pixelated photodetector each pixel has an area sufficient to collect the majority of the energy of the particle-beam interaction signal; wherein the pixelated photodetector produces an electric signal characteristic of the number and/or size of the particles detected.
 13. A system for detecting particles in a fluid, the system comprising: a flow cell for flowing a fluid containing particles along a flow direction through a beam of electromagnetic radiation, an optical source in optical communication with the flow cell for providing the beam of electromagnetic radiation; a focusing system in optical communication with the optical source for focusing said beam of electromagnetic radiation to generate an area of high radiation density within said flow cell; and an optical collection system for collecting and directing at least a portion of electromagnetic radiation onto a pixelated photodetector, wherein for at least a portion of the pixels of said pixelated photodetector each pixel has an area corresponding to a spatial extent of the particle beam interaction signal within the beam; wherein the pixelated photodetector produces an electric signal characteristic of the number and/or size of the particles detected.
 14. A system for detecting particles in a fluid, the system comprising: a flow cell for flowing a fluid containing particles along a flow direction through a beam of electromagnetic radiation, an optical source in optical communication with the flow cell for providing the beam of electromagnetic radiation; a focusing system in optical communication with the optical source for focusing said beam of electromagnetic radiation to generate an area of high radiation density within said flow cell; and an optical collection system for collecting and directing at least a portion of electromagnetic radiation onto a pixelated photodetector, wherein said optical collection system recollimates or focuses said beam of electromagnetic radiation; wherein for at least portion of the pixels of the pixelated photodetector each pixel has an area corresponding to a spatial extent of the particle beam interaction signal within the beam; wherein the photodetector produces an electric signal characteristic of the number and/or size of the particles detected.
 15. The system of claim 12, wherein each pixel has a width sufficient to collect the majority of the energy of the particle-beam interaction signal.
 16. The system of claim 12, wherein each pixel has a width independently selected from the range of 10 to 500 microns.
 17. The system of claim 12, wherein each pixel has a width sufficient to collect 80% or greater of the energy of the particle-beam interaction signal.
 18. The system of claim 12, wherein the at least a portion of the pixels of said pixelated photodetector each has an area matched to at least 75% of the spatial extent of the particle beam interaction signal within the beam. 19-23. (canceled)
 24. The system of claim 12, wherein the optical collection system comprises: an on-axis optical collection system for collecting and directing at least a portion of electromagnetic radiation onto a photodetector, thereby generating an on-axis signal; and a side scatter detector in optical communication with said flow cell position to receive off-axis scattered light, thereby generating a side scattered signal; wherein comparison of the side scattered signal to the on-axis signal distinguishes between biological end non-biological particles.
 25. (canceled)
 26. The system of claim 12, wherein each detector element of the pixelated photodetector produces an electric signal characteristic of the number and/or size of the particles detected and said photodetector characterizes said particles based on a differential signal generated from the detector element signals, the system comprising: a coprocessor configured to receive said detector element signals and characterize whether said particles have a lower or higher refractive index than said fluid.
 27. The system of claim 26, wherein the processor is configured to characterize the refractive index of the particle via the sequence, order and/or position of a dark fringe and a bright fringe as a function of time during the trajectory of the particle through the beam.
 28. The system of claim 27, wherein the processor is configured to characterize said particles as a metal or a non-metal via the refractive index of the particle.
 29. The system of claim 12 comprising: an adjuster operably connected to said photodetector or to said focusing system; wherein said adjuster moves said photodetector or alters said focusing system to balance the differential detection of the at least two elements of the photodetector.
 30. (canceled)
 31. (canceled)
 32. The system of claim 29, wherein the adjustor is controlled via closed loop feedback control.
 33. The system of claim 29, wherein said adjuster is operably connected to said photodetector and translates, moves, rotates or tilts said photodetector.
 34. The system of claim 29, wherein said adjuster is a steering mirror or a lens operably connected to said focusing system and adjusts a path of said beam of electromagnetic radiation.
 35. (canceled)
 36. The optical collection system of claim 35 further comprising an imager, wherein said beam of electromagnetic radiation may be directed toward said imager and said imager provides feedback to said adjuster in a closed loop on optimal optical beam power density, optimal beam spot size, optimal area of high radiation density in said flow cell or any combination thereof.
 37. The system of claim 12, wherein said beam of electromagnetic radiation is a Gaussian beam.
 38. The system of claim 12, wherein said beam of electromagnetic radiation is a structured beam.
 39. The system of claim 12, wherein said beam of electromagnetic radiation is a dark beam.
 40. The system of claim 12, wherein said beam of electromagnetic radiation is an anamorphic beam in a top hat configuration.
 41. The system of claim 12, wherein said photodetector comprises at least two detector elements and characterizes said particles based on a differential signal from individual signals from each detector element indicative of said particles. 42-45. (canceled)
 46. The system of claim 12, wherein said focusing system directs said beam of electromagnetic radiation through said flow cell at least twice and said particles in said flow cell interact with a different portion of said beam on each individual pass through said flow cell.
 47. (canceled)
 48. The system of claim 12, wherein said focusing system comprises a halfwave plate, a quarter wave plate or both for altering a polarization state of said beam. 49-58. (canceled)
 59. The system of claim 12, comprising: a translator operably connected to said flow cell for translating said flow cell closer to and further away from said focusing system such that said area of high radiation density changes position in said flow cell.
 60. The system of claim 59, wherein said translator is an oscillator.
 61. The system of claim 60, wherein said oscillator is a piezoelectric oscillator.
 62. The system of claim 24, wherein the on-axis optical collection system is disposed at a scattering angle that is within 1 degree of zero degrees relative to the beam of electromagnetic radiation. 